SPAK and OSR1 kinases role in the regulation of ion transport in Kir2.1 (KCNJ2), CreaT (SLC6A8) and NaPi-IIb (SLC34A2)

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dc.contributor.advisor Lang, Florian (Prof. Dr. med. Dr. h.c.) Fezai, Myriam 2018-04-05T09:32:24Z 2018-04-05T09:32:24Z 2018-04-05
dc.identifier.other 507419626 de_DE
dc.identifier.uri de_DE
dc.description.abstract The regulation of ion transport involves several kinases including SPAK (SPS1-related proline/alanine-rich kinase) and OSR1 (oxidative stress-responsive kinase 1), which are controlled by WNK (with-no-K[Lys]) kinases. The present study investigates whether SPAK and/or OSR1 participate in the modulation of the K+ channel Kir2.1 (KCNJ2), the creatine transporter CreaT (SLC6A8) and the phosphate transporter NaPi-IIb (SLC34A2). The first part of the study focuses on Kir2.1, namely: the inwardly rectifying K+ channel, which is expressed in brain, heart and skeletal muscle. Kir2.1 maintains the resting membrane potential and is determinant for cell volume regulation, preventing cell shrinkage. In addition kir2.1 participates in the spatial K+ buffering in neurons and contributes hence to neuro-excitability. The inwardly rectifying K+ current in Kir2.1 co-expressing Xenopus laevis oocytes is significantly increased in the presence of SPAK and OSR1 respectively; as a result of the enhanced channel protein abundance on the cell surface. Moreover, treatment with Brefeldin A abolishes the stimulating effect of SPAK or OSR1 on Kir2.1 indicating that the kinases effect involves in the protein trafficking pathway towards the plasma membrane. Further investigation regarding the kinases mutations show that the constitutively active mutants T233ESPAK and T185EOSR1 mimic the effect of the corresponding wild type kinase, while neither the WNK insensitive T233ASPAK and T185AOSR1, nor the catalytically inactive D212ASPAK and D164AOSR1 enhanced kir2.1 activity. The second part of the investigation concerns CreaT, i.e. the Na+ and Cl- coupled creatine transporter. CreaT carrier is expressed in cells with high energy demand such as brain, heart, intestine, retina and skeletal muscle, where it provides creatine needed for energy. The conversion of creatine to phosphocreatine by creatine kinase is important for maintaining ATP in the cells. Creatine transporter is additionally involved in mental retardation, seizure and intellectual disability. The co-expression of SPAK or OSR1 with CreaT decreases significantly the electrogenic creatine transport. Furthermore, the maximal creatine-induced inward current indicating the rate of transport is down-regulated by SPAK or OSR1, whereas the affinity of CreaT carrier to creatine is not affected. Besides the results of wild type kinases, the respective active mutants namely the constitutively active T233ESPAK and T185EOSR1 do negatively regulate CreaT, however, the catalytically inactive mutants D212ASPAK and D164AOSR1 as well as the WNK insensitive T233ASPAK do not, while T185AOSR1 tend to reduce creatine current. The last part of the study, explores the regulation of the Na+ coupled phosphate transporter NaPi-IIb, which ensures phosphate up-take in the intestine and tumor cells. The carrier is stimulated by SPAK and OSR1. Both kinases increase not only the maximal phosphate inward current and thus the transport rate, but also enhance significantly the affinity of NaPi-IIb to phosphate. Moreover, the simultaneous expression of SPAK and OSR1 do not show an additive effect. Furthermore, the constitutively active mutants T233ESPAK and T185EOSR1 display same effect as the wild type kinase respectively on NaPi-IIb activity. This effect is observed neither with the inactive mutants WNK insensitive T233ASPAK and T185AOSR1 nor with the catalytically inactive D212ASPAK and D164AOSR1. In conclusion, both kinases SPAK and OSR1 are powerful regulators of Kir2.1, NaPi-IIb and CreaT, effects may be involved in the cell volume regulation and excitability. en
dc.language.iso en de_DE
dc.publisher Universität Tübingen de_DE
dc.rights ubt-podok de_DE
dc.rights.uri de_DE
dc.rights.uri en
dc.subject.classification Ionenkanal , Elektrophysiologie de_DE
dc.subject.ddc 570 de_DE
dc.subject.other SPAK en
dc.subject.other OSR1 en
dc.subject.other kinase en
dc.subject.other Kir2.1 channel en
dc.subject.other CreaT and NaPi-IIb transporters en
dc.title SPAK and OSR1 kinases role in the regulation of ion transport in Kir2.1 (KCNJ2), CreaT (SLC6A8) and NaPi-IIb (SLC34A2) en
dc.type PhDThesis de_DE
dcterms.dateAccepted 2018-03-16
utue.publikation.fachbereich Medizin de_DE
utue.publikation.fakultaet 4 Medizinische Fakultät de_DE


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